Agarose is the most universally utilized chromatographic gel matrix for the separation of biopolymers in such applications as gel filtration, ion exchange chromatography, affinity chromatography, dye ligand chromatography, and hydrophobic interaction chromatography. The wide applicability of agarose is based on its neutrality and hydrophilicity, the ease with which it can be derivatised and cross-linked and its porosity and pH-stability.
Until recently, agarose has been used exclusively for conventional low-pressure chromatography. However, it has been shown that agarose can be used also for High Performance Liquid Chromatography (HPLC) which makes the application range for agarose still wider.
A decisive requirement of a bed material for HPLC is that it can tolerate relatively high pressures without appreciable deformation. Some compressibility may, however, be desirable since the void volume then decreases more than the total volume, which leads to an increase in resolution.
Cross-linked agarose gels, which are still permeable to proteins and particles, fulfill this requirement.
Divinylsulfone (DVS) belongs to the group of cross-linking agents which permit high flow rates.
From J. Chromatogr. 103 (1975) 49-62 it is known to cross-link agarose gels with DVS but in practice it has been found that such gels exhibit the following drawbacks that limit their usefulness as chromatographic bed materials:
(1) Blue Dextran used as a void volume marker was irreversibly adsorbed. PA0 (2) Some model proteins were more or less adsorbed (hemoglobin irreversibly). PA0 (3) The reproducibility in the cross-linking experiments was not satisfactory, since the maximum flow rate obtainable varied from batch to batch and was sometimes relatively low. PA0 (4) In the above article it was stated that the DVS-cross-linked gels were not stable above pH 9.